Efficacy, safety and immunological profile of combining rituximab with belimumab for adults with persistent or chronic immune thrombocytopenia: results from a prospective phase 2b trial

Abstract

B-cell activating factor may be involved in the failure of B-cell depleting therapy with rituximab in immune thrombocytopenia (ITP) by promoting the emergence of splenic long-lived plasma cells. From results obtained in mouse models, we hypothesized that combining rituximab with sequential injections of belimumab could increase the rate of response at one year in patients with persistent or chronic ITP by preventing the emergence of these long-lived plasma cells. The study was a single-center, single arm, prospective phase 2b trial (RITUX-PLUS, NCT03154385) investigating the safety and efficacy of rituximab given at a fixed dose of 1,000 mg, two weeks apart, combined with five infusions of belimumab, 10 mg/kg at week 0 (W0)+2 days, W2+2 days, W4, W8 and W12 for adults with primary persistent or chronic ITP. The primary endpoint was the total number of patients achieving an overall response (complete response + response) at W52 according to a standard definition. In total, 15 non-splenectomized adults, nine (60%) with persistent IPT and six (40%) with chronic ITP, were included. No severe adverse event, infection, or severe hypogammaglobulinemia was observed. Thirteen patients achieved an initial overall response. At W52, 12 (80%) patients achieved an overall response, including ten (66.7%) with complete response. When compared with a cohort of patients receiving rituximab alone, the kinetics of B-cell repopulation appeared similar, but the number of circulating T follicular helper cells was significantly decreased with belimumab combination therapy. Combining rituximab and belimumab seems a promising strategy in ITP, with high efficacy and acceptable safety.

Figures

Figure 1.

Serum level of total γ-globulins and immunoglobulin isotypes (IgG, IgA, IgM) during the study of rituximab and belimumab combined. Serum level of total γ-globulins (A) and IgG (B), IgM (C), and IgA (D) were assessed by nephelometry at week 12 (W12), W24, W36, W52. Dotted line represents normal threshold for each isotype. *P<0.05, **P<0.01, ***P<0.001; ns: not significant.

Figure 2.

Efficacy of rituximab and belimumab combination in adults with persistent and chronic immune thrombocytopenia. (A) Outcome at week 4 (W4), W12, W24, W36, and W52 according to international recommendations. Complete response (CR) was defined by a platelet count >100x109/L and response (R) by a platelet count 30-100x109/L with at least a 2-fold increase from baseline. Non-responders (NR) are labeled in red; platelet counts were censured when an ITP-directed therapy was started. (B) Evolution of platelet count for each patient during the study. ITP: immune thrombocytopenia; NR: no response.

Figure 3.

B-cell activating factor and B-cell subsets in immune thrombocytopenia patients receiving the combination therapy or rituximab alone. (A) B-cell activating factor (BAFF) concentrations were assessed by enzyme-linked immunosorbent assay in serum of patients receiving rituximab and belimumab (in blue) or rituximab alone (in red) at week 0 (W0), W12, W24, W36 and W52. Data are mean ± standard error of the mean (pg/mL). (B) Gating strategy of B-cell subpopulations. Single lymphoid cells on peripheral blood mononuclear cells (PBMC) were gated by using scatter parameters, and dead cells were eliminated by using zombie violet. Plasmablasts/plasma cells (PB/PC) were defined as CD27hiCD38hi cells among CD3-CD14-CD16- cells. After excluding CD3/CD14/CD16-positive cells and PB/PC from the CD19+ gate, B-cell subsets were separated according to their expression of CD27 and IgD and defined as memory B cells (CD27+IgD-), CD27+IgD+ B cells, and naïve B cells (CD27–IgD+). Transitional B cells were defined as CD38hiCD24hiCD10+ cells among naïve B cells. (C) Circulating B-cell subset count per million PBMC at W0, W4, W12, W24, W36 and W52. ****P<0.0001

Figure 4.

Rituximab and belimumab combination affects activated circulating T- follicular helper cells. (A) Gating strategy for circulating T- follicular helper cells (cTfh) cells. After gating on CD4+CD45RA– memory T-CD4+ cells in whole blood, cTfh cells were defined as CXCR5+PD1+, and activated cTfh cells as CXCR5+PD1+ICOS+. (B) Percentages of cTfh and (C) activated Tfh cells at week 0 (W0), W4, W12, W24, W36 and W52 in patients receiving rituximab and belimumab or rituximab alone. *P<0.05, **P<0.01.